This proposal describes a 5-year research plan focusing on the proximal signaling complexes that are formed downstream of natural killer (NK) cell activating receptors and how signaling pathways leading from these complexes affect NK cell effector function and development. NK cells are innate immune cells that defend the host from intracellular pathogens and tumors. Regulation of NK cell activation involves the expression of activating receptors that are finely counterbalanced by inhibitory MHC class I-binding receptors (e.g., Ly49 family members), which allow NK cells to achieve self tolerance. How the precise signaling pathways leading from NK cell activating receptors contribute to mature NK cell effector function and inhibitory receptor acquisition during development are not well understood. Signaling through activating immunoreceptors relies on the formation of a proximal multimolecular complex nucleated by the adaptor protein SH2 containing leukocyte protein of 76kD (SLP-76). Our preliminary data support an important role for SLP-76 in activating receptor-mediated NK cell effector signal transduction, which is critical for NK cell effector function and Ly49 receptor acquisition. In addition, our data suggest that NK cells also use an alternative mechanism that may be different from other hematopoietic cell types to localize SLP-76 to the plasma membrane and to initiate signal transduction. This alternative signal transduction pathway leading from SLP-76 may be important during NK cell development for the acquisition of Ly49 receptors. In this proposal, we hypothesize that SLP-76 mediates multiple signal transduction pathways downstream of NK cell activating receptors, which play differential roles in NK cell effector function and in the acquisition of Ly49 receptors. Our research plan starts with an examination of how proximal signaling complexes involving SLP-76 are formed in NK cells downstream of activating receptors. We will then examine how these signal transduction pathways contribute to the different NK cell effector functions and to NK cell acquisition of Ly49 receptors. The experiments described in this proposal will yield insight into the precise signal transduction pathways downstream of activating receptors in NK cells, which is essential for our understanding of how NK cells are activated in the periphery and achieve self tolerance through expression of Ly49 receptors. This information will allow us to devise novel strategies to modulate NK cell responses in multiple disease settings. Our expertise in signal transduction and NK cell biology together with our expert consultants (Drs. Jordan Orange, Tobias Baumgart, and Yoji Shimizu), and the quality of the research facilities at the University of Pennsylvania Medical Center comprise an ideal environment in which to conduct this proposed research plan. Thus, we believe that we are uniquely equipped with the tools/reagents and expertise to investigate this important aspect of NK cell biology.